The general field of this invention is therapeutic agents for combating viral infections. In particular, the present invention relates to the treatment of patients infected with human immunodeficiency virus (HIV) which causes Acquired Immuno Deficiency Syndrome (AIDS).
Human immunodeficiency virus (HIV) has been determined to be the etiologic agent for acquired immunodeficiency syndrome (AIDS). The past decade has seen an unprecedented massive, worldwide research effort to discover effective therapies for HIV/AIDS costing billions of dollars that has only yielded limited success. Current therapeutic agents used during the treatment of chronic phase of HIV infection have not been particularly efficacious and are, at best, only able to slow the progress of infection. For example, several drugs have been identified which inhibit the replication of HIV in vitro (Haseltine, W. A., J. Acquir. Immune Def. Syndr., 2:311-324, 1989) and nucleotide chain elongation inhibitors such as 3-azidothymidine (AZT) have received widespread acceptance for clinical use. However, the clinical efficacy of drugs like AZT has proven rather limited, and their use is further restricted by toxicity factors, and drug-resistant forms of the virus. Montefiori (U.S. Pat. No. 5,057,325), discovered that melanin produced by chemical oxidation when applied to HIV-infected cells in vitro exhibited anti-HIV activity.
Melanin is an omnibus term that describes a large family of natural and synthetic phenolic-quinonoid pigments of diverse origin and chemical nature. Natural melanins are generally differentiated by their origin; for example, bovine eye melanin, melanoma melanin and sepia melanin. They usually occur in the form of granular particles and are secretory products of pigment-producing cells, the melanocytes. Synthetic melanins are named after the compound from which they were prepared via chemical or enzymatic oxidation (e.g., d- and 1-dopa., or 5,6-dihydroxyindole catechol melanin). In addition, melanins are classified according to their chemical structure into the insoluble black eumelanins (poly-5,6-indole quinones) and the alkali soluble red phaeomelanins (polydihydrobenzothiazines). The study of melanins has led to the discovery of a number of biosynthetic pathways. For example, melanins can be produced by the oxidation of its precursors such as 1-dopa or tyrosine by the melanin-synthesizing enzyme, tyrosinase. Alternatively, melanin can be prepared chemically by the auto-oxidation of 1-dopa or other substrates to melanin, in the presence of atmospheric oxygen (Wilczok et al., Arch Biochem. Biophys. 23:257, 1984). Additionally, melanin can be prepared by a variety of electrochemical and photochemical methods from which individual steps of the melanization processes are identified and characterized. See, Crippa, et al., (1989), supra. In this invention, melanins are produced in either aqueous or organic solutions, using the tyrosinase-mediated polymerization of tyrosine and tyrosine dipeptides and mixtures of both (della-Cioppa, et al., op. cit.). Tyrosine dipeptides have been previously used to produce melanins with tyrosinase; however, prior to the present invention, no one had recognized that such compounds may possess anti-viral activity. Thus, the presently disclosed anti-viral functions and modifications were never investigated (Yasunobu, et al., 1959).
Individuals infected with HIV (HIV-positive) may not develop symptoms of AIDS for years, but eventually the vast majority of HIV-positive patients develop AIDS. AIDS patients suffer from an overwhelming assault of diseases and have a poor prognosis for long term survival even when subjected to aggressive therapeutic regimens. Consequently, there is a compelling need to develop an effective anti-HIV therapy to either eliminate or arrest the progression of HIV infection and to prevent or treat AIDS.
To achieve this end, a first aspect of the present invention relates to a novel method for arresting HIV replication as a therapy for persons exposed to the HIV virus or testing HIV-positive, and for AIDS patients by administering an effective dose of melanin derived from the controlled enzymatic action of tyrosinase in conjunction with specific substrates or mixtures of substrates.
A second aspect of the invention relates to methods for the chemical polysulfation of enzymatically or non-enzymatically produced melanin to increase antiviral activity, as well as the sulfated or polysulfated melanin products of such methods.
A third aspect of the invention relates to methods for the chemical polyhalogenation of melanin with fluorine, chlorine, bromine and/or iodine to increase antiviral activity, as well as the halogenated melanin products of such methods.
A fourth aspect of the invention relates to the therapeutic use of melanins that have been modified by any combination of polysulfation, polysulfonation, and/or polyhalogenation.
A fifth aspect of the invention relates to melanins produced by enzymatic synthesis using defined substrates, and modified forms thereof, that have been purified by precipitation by reducing the pH of a melanin containing solution below about 7.0, and preferably below about pH 3.0, with a preferred pH of about 1.5.
A sixth aspect of the invention relates to melanin produced by enzymatic synthesis wherein said melanin is oxidized by hydrogen peroxide or another suitable chemical oxidizer.
A seventh aspect of the invention relates to melanin produced by enzymatic synthesis wherein said melanin is further purified by passage through a suitable molecular sieve to obtain melanin having a molecular size greater than or equal to about 10,000 daltons.
An eighth aspect of the invention relates to melanin produced by either chemical or enzymatic synthesis wherein said melanin is further modified to have chemical subgroups selected from the group consisting of sulfonyl, sulfate and halogen chemical groups, or any mixture thereof.